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Biomedical subjects

P Moretti

Publications and source records attributed to P Moretti.

At least 37 records · Page 2Linked to original sources
PubMed1994

Evidence that a complex of SIR proteins interacts with the silencer and telomere-binding protein RAP1.

The maintenance of transcriptional silencing at HM mating-type loci and telomeres in yeast requires the SIR2, SIR3, and SIR4 proteins, none of which appear to be DNA-binding proteins. Here we show that SIR3 and SIR4 interact with a carboxy-terminal domain of the silencer, telomere, and UAS-binding protein RAP1. We identified SIR3 and SIR4 in a two-hybrid screen for RAP1-interacting factors and showed that SIR3 interacts both with itself and with SIR4. The interaction between RAP1 and SIR3 can be observed in vitro in the absence of other yeast proteins. Consistent with the notion that native SIR proteins interact with the RAP1 carboxyl terminus, we show that mutation of the endogenous SIR3 and SIR4 genes increases transcriptional activation by LexA/RAP1 hybrids. To test the importance of the RAP1-SIR3 interaction for silencing, we identified mutations in the RAP1 carboxyl terminus that either diminish or abolish this interaction. When introduced into the native RAP1 protein, these mutations cause corresponding defects in silencing at both HMR and telomeres. We propose that RAP1 acts in the initiation of transcriptional silencing by recruiting a complex of SIR proteins to the chromosome via protein-protein interactions. These data are consistent with a model in which SIR3 and SIR4 play a structural role in the maintenance of silent chromatin and indicate that their action is initiated at the silencer itself.

Bacterial Proteins↗
PubMed1994

Identification of homeobox genes expressed in human haemopoietic progenitor cells.

Homeodomain (HD)-containing proteins have been shown to regulate cellular commitment and differentiation in fungal, invertebrate and vertebrate systems. Bone marrow cells synthesizing the CD34 antigen are a complex mix of early, stem and progenitor cells at various stages of commitment to the many haemopoietic lineages. Here, we report the cloning and sequencing of 31 homeobox (HB) sequences, identified using degenerate oligodeoxyribonucleotide primers, in a polymerase chain reaction with cDNA derived from a purified CD34+ population of human haemopoietic cells. Of these sequences, 16 correspond to previously identified genes, and 13 are located within the HOX A, B and C clusters. Ten of the clones most likely represent human homologues of genes identified previously in other species. Five of the clones reported here represent novel HD sequences. The identification of five new genes using a subclass-specific 5' primer, designed from the engrailed and Xanf1 sequences, suggests that there still remain several uncharacterized HB genes in the human genome. Haemopoietic cells purified on the basis of CD34 antigen synthesis are a rich source of regulatory genes consistent with their ability to differentiate into diverse haemopoietic cell types.

Amino Acid Sequence↗
PubMed1994

A mutation of the common receptor subunit for interleukin-3 (IL-3), granulocyte-macrophage colony-stimulating factor, and IL-5 that leads to ligand independence and tumorigenicity.

The cytokines interleukin-3, interleukin-5, and granulocyte-macrophage colony-stimulating factor bind with high affinity to a receptor complex that contains a ligand-specific alpha-chain and a common beta-chain, h beta c. We report here the isolation of a mutant form of h beta c, from growth factor-independent cells, that arose spontaneously after infection of a murine factor-dependent hematopoietic cell line (FDC-P1) with a retroviral h beta c expression construct. Analysis of this h beta c mutation shows that a small (37 amino acid) duplication of extracellular sequence that includes two conserved sequence motifs is sufficient to confer ligand-independent growth on these cells and lead to tumourigenicity. Because this is a conserved region in the cytokine receptor superfamily, our results suggest that the large family of cytokine receptors has the capacity to become oncogenically active.

Amino Acid Sequence↗
PubMed1994

Two contiguous residues in human interleukin-3, Asp21 and Glu22, selectively interact with the alpha- and beta-chains of its receptor and participate in function.

We have previously reported that the predicted first helix of human interleukin (IL)-3 contains a hydrophilic region encompassing residues Asp21, Glu22, and Thr25 that is crucial for biological activity and IL-3 receptor binding. Using single amino acid substitution mutagenesis, we have now determined that Asp21 and Glu22, but not Thr25, were crucial for full IL-3 activity. Mutant D21R was 30-fold less potent than wild type IL-3 in the stimulation of biological activity. It also exhibited a similar reduction in its ability to bind to the cloned high affinity IL-3 receptor complex (alpha- and beta-chains) or to the receptor alpha-chain alone, indicating that residue 21 is involved in contacts with the alpha-chain. Mutant E22R was approximately 20,000-fold less potent than wild type IL-3 in the stimulation of biological activity and in binding to the IL-3 receptor high affinity complex. However, the binding of E22R to the IL-3 receptor alpha-chain alone was similar to that of wild type IL-3, suggesting that this mutant was defective in interactions with the receptor beta-chain. These results show that two contiguous residues in the N-terminal region of IL-3 mediate binding to the two different chains of the IL-3 receptor and emphasize the functional significance of the conserved Glu in the first helix of the IL-3, granulocyte-macrophage colony-stimulating factor, and IL-5 cytokine subfamily.

Amino Acid Sequence↗
PubMed1994

Structure and regulation of the mouse cardiac troponin I gene.

The gene coding for mouse cardiac troponin I (TnI) has been cloned and sequenced. The cardiac TnI gene contains 8 exons and has an exon-intron organization similar to the quail fast skeletal TnI gene except for the region of exons 1-3, which is highly divergent. Comparative analysis suggests that cardiac TnI exon 1 corresponds to fast TnI exons 1 and 2 and that cardiac exon 3, which codes for most of the cardiac-specific amino-terminal extension and has no counterpart in the fast gene, evolved by exon insertion/deletion. The amino acid sequence of cardiac TnI exon 4 shows limited homology (36% identity) with fast TnI exon 4 but is remarkably similar (79% identity) to the corresponding sequence of slow TnI, possibly reflecting an isoform-specific TnC-binding site. The cardiac TnI gene is one of the very few contractile protein genes expressed exclusively in cardiac muscle. To identify the regulatory sequences responsible for the cardiac-specific expression of this gene we transfected cultured cardiac and skeletal muscle cells with fragments up to 4.0 kilobases of the 5'-flanking region linked to a reporter gene. Deletion analysis reveals four major regions in the 5'-flanking sequence, a minimal promoter region, which directs expression at low level in cardiac and skeletal muscle cells, and two upstream cardiac-specific positive regions separated by a negative region.

Amino Acid Sequence↗
PubMed1993

Transient topographical amnesia: a case report.

We describe the case of a 56 year old female, with history of migraine since adolescence, who experienced two episodes of transient topographical disorientation in the absence of intellectual deterioration or evident focal cerebral lesions.

Acute Disease↗
PubMed1993

Type 2X-myosin heavy chain is coded by a muscle fiber type-specific and developmentally regulated gene.

We have previously reported the identification of a distinct myosin heavy chain (MyHC) isoform in a major subpopulation of rat skeletal muscle fibers, referred to as 2X fibers (Schiaffino, S., L. Gorza, S. Sartore, L. Saggin, M. Vianello, K. Gundersen, and T. Lømo. 1989. J. Muscle Res. Cell Motil. 10:197-205). However, it was not known whether 2X-MyHC is the product of posttranslational modification of other MyHCs or is coded by a distinct mRNA. We report here the isolation and characterization of cDNAs coding a MyHC isoform that is expressed in type 2X skeletal muscle fibers. 2X-MyHC transcripts differ from other MyHC transcripts in their restriction map and 3' end sequence and are thus derived from a distinct gene. In situ hybridization analyses show that 2X-MyHC transcripts are expressed at high levels in the diaphragm and fast hindlimb muscles and can be coexpressed either with 2B- or 2A-MyHC transcripts in a number of fibers. At the single fiber level the distribution of each MyHC mRNA closely matches that of the corresponding protein, determined by specific antibodies on serial sections. In hindlimb muscles 2X-, 2A-, and 2B-MyHC transcripts are first detected by postnatal day 2-5 and display from the earliest stages a distinct pattern of distribution in different muscles and different fibers. The emergence of type 2 MyHC isoforms thus defines a distinct neonatal phase of fiber type differentiation during muscle development. The functional significance of MyHC isoforms is discussed with particular reference to the velocity of shortening of skeletal muscle fibers.

Amino Acid Sequence↗
PubMed1993

[The clinical evaluation and immunological response of a new measles vaccine].

For a number of years the measles vaccine has been tabled to be included among the compulsory vaccines with the not impossible aim of wiping out this disease. In practice, however, there have been isolated attempts to impose mass vaccination but, given that it is not compulsory, there has been a certain degree of resistance to the vaccine. In an attempt to interpret the reasons for this resistance we have identified three main causes: cultural heritage by family doctors and sometimes pediatricians ("a disease which is important to have had"). Limited information with underestimation of damaging effects of disease overemphasis of side-effects of vaccine and, among these, in particular fever which 30-35% of vaccinated children present between day 8-10, and the possible allergic reaction (chicken embryos-antibiotics). Using the latter as a starting point, we assessed the efficacy and tolerability of the Moraten Berna vaccine on 165 children aged between 10 months and 8 years. This vaccine is prepared using the Edmonston Zagreb strain cultured in diploid human cells, and does not therefore contain heterologous proteins, seroconversion was assessed in 54 of the vaccinated children (aged 10 months--3 years) by blood sample collected following an interval of between 40 and 120 days after vaccination. Anti-measles antibody titering was assayed using a neutralisation test on plates of cellular monostrate by assessing the titer (starting dilution of serum in test: 1:8). Seroconversion, as shown in table I, was obtained for all vaccinated subjects who presented a mean antibody titer of 916 mUI/ml.(ABSTRACT TRUNCATED AT 250 WORDS)

Antibodies, Viral↗
PubMed1992

A novel endothelial tyrosine kinase cDNA homologous to platelet-derived growth factor receptor cDNA.

Degenerate oligonucleotide primers complementary to the highly conserved subdomains III and VIII of subclass III tyrosine kinase receptors (TKr-III) were utilized to amplify rat aortic cDNA by polymerase chain reaction. Most of the cloned DNA products were rat platelet-derived growth factor receptor beta and macrophage-colony stimulating growth factor receptor cDNAs. Screening of the clones with probes coding for the receptor-specific kinase insert domain allowed the identification of a novel putative TKr-III cDNA, which hybridized with a approximately 6.1 kb mRNA with a distinctive tissue distribution. In situ hybridization on rat tissues and Northern analysis of cultured cells indicate that endothelial cells express a novel putative TKr-III mRNA.

Amino Acid Sequence↗
PubMed1992

Mini-Mental State Examination (MMSE): sensitivity in an Italian sample of patients with dementia.

The sensitivity of the Mini-Mental State Examination (MMSE) was assessed in a sample of patients with dementia of the Alzheimer type or vascular dementia. The MMSE identified the majority of pts with diffuse cognitive impairment but did not discriminate between the two types of dementia. If failed to detect mild deterioration or forms in which only some cognitive functions were impaired. The test is therefore not sufficient for distinguishing deteriorated from non deteriorated pts, although it is still useful in mass screening or for a quick assessment of deterioration in the course of clinical neurological examination.

Alzheimer Disease↗
PubMed1992

Supernova (spno), a new maternal mutant producing variable-sized cleavage nuclei in Drosophila.

This paper describes a new recessive maternal lethal which disrupts normal nuclear division and migration during cleavage in Drosophila. We have named this gene locus supernova. Deletion mapping and in situ hybridization have located this gene to 88 F9/89 A1 on the polytene chromosome map. The terminal mutant phenotype is characterized by the presence of many variable-sized nuclei scattered throughout the cytoplasm of the unhatched egg. Following fertilization, the initial cleavage divisions appear delayed and are often accompanied by the formation of ring-like association of chromosomes and/or chromosome bridges. Although the polymerization of tubulin into spindles occurs during the initial cleavage divisions, there appears to be both a spatial and temporal uncoupling of DNA replication from the formation and proper functioning of spindles. Eventually no functional spindles are formed, but nuclei continue to increase in size and number with increasing age of the embryo following fertilization.

Animals↗
PubMed1991

Emergence of the mature myosin phenotype in the rat diaphragm muscle.

Immunohistochemical analysis of myosin heavy chain (MHC) isoform expression in perinatal and adult rat diaphragm muscles was performed with antibodies which permitted the identification of all known MHC isoforms found in typical rat muscles. Isoform switching, leading to the emergence of the adult phenotype, was more complex than had been previously described. As many as four isoforms could be coexpressed in a single myofiber. Elimination of developmental isoforms did not usually result in the myofiber immediately achieving its adult phenotype. Activation of genes for specific adult isoforms might be delayed to puberty. For example, two of the three fast MHCs, MHC2X and MHC2A appeared perinatally, while MHC2B did not appear until 30 days postnatal. By Day 60 this isoform was present in approximately 27% of the myofibers, but in most myofibers expression of this isoform was transient (i.e., at Day greater than or equal to 115, less than 4% of the myofibers expressed MHC2B). Fibers which contained MHC beta/slow during the late fetal and early neonatal period coexpressed MHCemb. A marked increase in the frequency of fibers containing MHC beta/slow occurred between 4 and 21 days postnatal. These slow fibers arose from a population of myofibers which expressed MHCemb and MHCneo during their development, and they accounted for the majority of slow fibers found in the adult diaphragm. The adult myosin phenotype of the diaphragm myofibers (as determined with immunocytochemistry, and 5% SDS-PAGE) was not achieved until the rat was greater than or equal to 115 days old.

Age Factors↗
PubMed1991

Developmental expression of rat cardiac troponin I mRNA.

We have isolated and sequenced a full-length cDNA clone of rat cardiac troponin I (TnI). The amino acid sequence of rat cardiac TnI is highly similar to that of other mammalian species in the portion of the molecule (residues 33-210) that is also homologous to skeletal muscle TnI isoforms. In contrast, a lower degree of similarity is present in the cardiac TnI-specific amino terminal extension (residues 1-32). This region contains a conserved serine residue that has been shown to be selectively phosphorylated by cAMP-dependent protein kinase. Cardiac TnI mRNA is weakly expressed in the 18-day fetal heart and accumulates in neonatal and postnatal stages. No difference can be demonstrated between TnI mRNAs present in fetal and postnatal heart by RNAase protection assays. The fetal and neonatal, but not the adult heart, contain significant amounts of slow skeletal TnI transcripts, detected by oligonucleotide probes specific for the 5'- and 3'-untranslated regions of slow skeletal TnI mRNA. In situ hybridization studies show that cardiac and slow skeletal TnI mRNAs are coexpressed in the rat heart from embryonic day 11 throughout fetal and perinatal stages. Changes in troponin isoform expression during development may be responsible for the difference in calcium sensitivity and in the response to beta-adrenergic stimulation between fetal and adult heart.

Amino Acid Sequence↗
PubMed1990

Electrophoretic separation and immunological identification of type 2X myosin heavy chain in rat skeletal muscle.

One slow and three fast myosin heavy chains have been described in typical skeletal muscles of the adult rat using immunocytochemical analysis. Electrophoretic isolation and immunochemical identification of these four isoforms has not been achieved. An electrophoretic procedure is described which, by altering the cross-linkage and polymerization kinetics of 5% polyacrylamide gels, allows resolution of these four distinct myosin heavy chains. Using specific monoclonal antibodies and double immunoblotting analysis, the identity and electrophoretic migration order of the myosin heavy chains was established to be: 2A less than 2X less than 2B less than beta/slow.

Animals↗
PubMed1990

Relationships between biotin and thymus morphology, and thymic and plasma peptides controlling DNA transcription.

Biotin-deficient rats show a slower growth rate and thymus involution. The amount of the thymic peptides controlling DNA template, based on thymus weight, is higher in deficient than control rats. No significant difference was noticed in the content of active peptides when evaluated for the rat. This observation suggests that involution of the thymus does not involve those cells which produce active peptides. Inhibition of RNA synthesis is the same for peptides extracted from normal and from biotin-deficient rat thymus. The amount of plasma peptides controlling DNA template per milliliter of plasma is higher and inhibition of activity on RNA synthesis is lower in deficient as compared to control rats. These effects are reversible following biotin administration.

Animals↗